toxB Gene on pO157 of Enterohemorrhagic Escherichia coli O157:H7 Is Required for Full Epithelial Cell Adherence Phenotype

Abstract

Adherence of enterohemorrhagic Escherichia coli (EHEC) to the intestinal epithelium is critical for initiation of a bacterial infection. An in vitro infection study previously indicated that EHEC bacteria initially adhere diffusely and then proliferate to develop MC, a process that is mediated by various secreted proteins, such as EspA, EspB, EspD, Tir, and intimin, as well as other putative adherence factors. In the present study, we investigated the role of a large 93-kb plasmid (pO157) in the adherence of O157:H7 (O157Sakai) and found the toxB gene to be involved in the full adherence phenotype. A pO157-cured strain of O157Sakai (O157Cu) developed microcolonies on Caco-2 cells; however, the number of microcolonies was lower than that of O157Sakai, as were the production and secretion levels of EspA, EspB, and Tir. Introduction of a mini-pO157 plasmid (pIC37) composed of the toxB and ori regions restored full adherence capacity to O157Cu, including production and secretion of the proteins. In contrast, introduction of a pO157 mutant possessing toxB::Km into O157Cu could not restore the full adherence phenotype. Expression of truncated versions of His-tagged ToxB also promoted EspB production and/or secretion by O157Cu. These results suggest that ToxB contributes to the adherence of EHEC to epithelial cells through promotion of the production and/or secretion of type III secreted proteins.

Quantitative Proteomic Analysis of Type III Secretome of Enteropathogenic Escherichia coli Reveals an Expanded Effector Repertoire for Attaching/Effacing Bacterial Pathogens

“…ToxB is encoded on the pO157 plasmid in EHEC O157:H7 strains (41). This suggests that lifA/efa1/toxB and the LEE (and its T3SS) are closely associated and that LifA/Efa1/ToxB are probably core effectors.…”

“…Their proposed functions in different A/E pathogens range from being a lymphotoxin and immune modulator (37,55), an adhesion/colonization factor (38,40,41), and a regulator of type III secretion (40,41,56), to being a modulator of Rho GTPases and intestinal epithelial-barrier function (52). These observations suggest that LifA/Efa1/ToxB have host cell binding activities and likely exert some of their functions from the bacterial side.…”

Mecanismos de virulencia de Escherichia coli enteropatógena

“…• Otros factores de virulencia de ECST: Adicionalmente, el plásmido pO157 de ECST codifica genes para catalasa-peroxidasa (katP) 56 , adhesina ToxB 57 , metaloproteasa dependiente de zinc (StcE) 58-61 y para la enterohemolisina (Ehx) 59 , cuyas funciones están principalmente enfocadas a mediar la unión a la célula hospedera, degradación de mucinas y glicoproteínas, regular mecanismos de inflamación y citotoxicidad, entre otras [56][57][58][59][60][61][62][63] .…”

Efa-1/LifA mediates intestinal colonization of calves by enterohaemorrhagic Escherichia coli O26 : H– in a manner independent of glycosyltransferase and cysteine protease motifs or effects on type III secretion

“…The reasons why efa-1/lifA mutations affect expression and secretion of type III secreted proteins in some EHEC strains but not others require further investigation. It is possible that functional redundancy may exist, whereby loss of Efa-1/LifA in some strains may be compensated for by other proteins that are similarly able to modulate type III secretion, such as ToxB (Tatsuno et al, 2001). Indeed, we observed that EHEC O26 strain 193 harbours toxB, whereas EHEC O111 strain E45035N and EHEC O5 strain S102-9 did not contain the gene as assessed by PCR with toxB-specific primers (Tozzoli et al, 2005; data not shown).…”

“…EHEC O157 : H7 strains also contain a homologue of Efa-1/LifA on their large plasmid (ToxB). ToxB promotes adherence of EHEC O157 : H7 to cultured cells and, as with efa-1/lifA in EHEC O5 : H-and O111 : H-, mutation of the gene impairs the expression and secretion of LEE4-encoded proteins at a post-transcriptional level (Tatsuno et al, 2001). However, ToxB was not required for intestinal colonization of calves and neither it, nor the truncated Efa-1, appears to confer lymphostatinlike activity (Abu-Median et al, 2006).…”